RU1774624C - 1-propyl-2-oxo-4-hydroxyquinoline-3-carboxylic acid diethylaminoethylamide hydrochloride showing anesthetic, antiarrhythmic, antioxidant, antimicrobial and fungicidal activity - Google Patents

Abstract

FIELD: organic chemistry. SUBSTANCE: product - 1-propyl-2-oxo-4-hydroxyquinoline-3-carboxylic acid diethylaminoethylamide hydrochloride, empirical formula is C₁₉H₂₇N₃O₃·HCl x HCl, m. p. is 160-162 C, yield is 84% . Reagent 1: 1-propyl-2-oxo-3-carbethoxy-4-hydroxyquinoline. Reagent 2: diethylaminoethylamine. Condition: boiling in methanol medium. Product is used in surgery. EFFECT: improved method of synthesis. 6 tbl

Description

I проявляющего анестезирующее, противоаритмическое, антиоксидантное, антимикробное и фунгицидное действие.The invention relates to the pharmaceutical industry and relates to biologically active substances, in particular 1-propyl-2-oxo-4-hydroxyquinoline-3-carboxylic acid diethylaminoethylamide hydrochloride of the formula I

I exhibiting anesthetic, antiarrhythmic, antioxidant, antimicrobial and fungicidal effects.

 These properties suggest the possibility of its use in medicine, in particular in surgery, as a complex drug.

CONHCH₂CH₂N(C₂H₅)₂·HCl
II проявляющий местноанестезирующую, антимикробную и противогрибковую активность.An analog in structure and action is 1-ethyl-2-oxo-4-hydroxyquinoline-3-carboxylic acid diethylaminoethylamide hydrochloride of formula II

CONHCH ₂ CH ₂ N (C ₂ H ₅ ) ₂ · HCl
II exhibiting local anesthetic, antimicrobial and antifungal activity.

 The analogue in action is the drug widely used in medical practice - lidocaine.

 The purpose of the invention is the search in a series of derivatives of 2-oxo-4-hydroxyquinoline-3-carboxylic acids of a new compound with higher anesthetic, antifungal and fungicidal activity.

 The goal is achieved by the described diethylaminoethylamide 1-propyl-2-oxo-4-hydroxyquinoline-3-carboxylic acid hydrochloride.

 PRI me R 1. Diethylaminoethylamide 1-propyl-2-oxo-4-hydroxyquinoline-3-carboxylic acid hydrochloride of the formula I.

To a solution of 2.75 g (0.01 mol) of 1-propyl-2-oxo-3-carbethoxy-4-hydroxyquinoline in 10 ml of methanol, 1.3 g (0.012 mol) of diethylaminoethylamine are added and refluxed for 5 hours. Cool 30 ml of ice water are added. The precipitate formed is filtered off, washed with water and dried. Then dissolved in 15 ml of methanol containing 0.011 mol of HCl. 5 ml of hexane are added and cooled. The precipitated crystals are filtered off, washed with hexane, and dried. Yield 3.20 g (84%); so pl. 160-162 ^about S.

 Found,%: C 59.62; H 7.50; N, 11.10; Cl 9.17.

C ₁₉ H ₂₇ N ₃ O ₃ ˙HCl.

 Calculated,%: C 59.76; H 7.39; N, 11.00; Cl 9.28.

PMR spectrum: 16.92 (1H, s, OH); 10.69 (1H, s. NH); 10.48 (1H, t, NH); 8.11 (1H, dd, H-5); 7.81 (1H, td, H-7); 7.66 (1H, d, H-8); 7.37 (1H, td, H-6); 4.21 (2H, q, NCH ₂ -C ₂ H ₅ ); 3.77 (2H, q, NHCH ₂ ); 3.23 (6H, m, N (CH ₂ ) ₃ ); 1.65 (2H, m, NCH ₂ CH ₂ CH ₂ ); 1.26 (6H, t, NCH ₂ CH ₃ x2); 0.97 (3H, t, CH ₂ CH ₂ CH ₃ ).

 PRI me R 2. 1-Propyl-2-oxo-3-carb-ethoxy-4-hydroxyquinoline.

To a solution of 2.07 g (0.01 mol) of N-propylanthranilic acid ethyl ester in 15 ml of acetone, 1.4 ml (0.01 mol) of triethylamine and 1.51 g (0.01 mol) of ethoxy maximonyl chloride are added. Leave for the night. Acetone is removed, a solution of 2.0 g KOH in 20 ml of water is added to the residue. Stirred for 1 h at 50 ^about C. Cool, acidify with HCl to pH 4. The residue was filtered off, washed with water, dried. Yield 2.09 g (76%); t. pl. 76-78 ^about (aqueous ethanol).

 Found,%: C 65.51; H 6.16; N, 5.14.

C ₁₅ H ₁₇ NO ₄ .

 Calculated,%: C 65.44; H 6.22; N, 5.09.

PMR spectrum: 13.07 (1H, s, OH); 8.05 (1H, dd, H-5); 7.76 (1H, td, H-7); 7.56 (1H, d, H-8); 7.29 (1H, td, H-6); 4.35 (2H, q, OCH ₂ ); 4.14 (2H, t, NCH ₂ ); 1.60 (2H, m, NCH ₂ CH ₂ ); 1.31 (3H, t, OCH ₂ CH ₃ ); 0.95 (3H, t, NCH ₂ CH ₂ CH ₃ ).

 The MPR spectra of the synthesized compounds were recorded on a Bruker WP-100 SY instrument (Germany), an operating frequency of 100 MHz, a DMSO-D6 solvent, chemical shifts are shown in the δ scale with respect to TMS.

PRI me R 3. The acute toxicity of compound 1 and comparison drugs was determined on white mice of both sexes weighing 20-25 g of 5 animals in a series with each dose. The test substances were dissolved in a 0.9% NaCl solution and administered intraperitoneally in a volume not exceeding 0.55 ml in a dose range of 40 to 150 mg / kg. Observations of animals were conducted for 14 days. LD ₅₀ (see table 1) was calculated by the Kerber method.

 PRI me R 4. The anesthetic activity of each compound under conditions of infiltration anesthesia was studied on 6 guinea pigs of red-black and white color, standardized by weight and gender, according to the Balbring method. The test compounds in the form of a 0.5% solution prepared in a 0.9% NaCl solution were administered intradermally in a volume of 0.25 ml. Anesthetic activity was assessed every 10 min by conducting a series of injections at the injection site of the test compound (5-6 needle injections). Anesthesia was considered adequate if the animals did not show signs of anxiety, there was no vocalization and reaction of the cardiovascular system (increased heart rate).

0,7
Результаты учитывали по отсутствию глубокой болевой чувствительности и двигательной активности в соответствующих группах мышц в области действия эпидуральной анестезии (см. табл.2).Example 5. The study of the anesthetic activity of compound F and lidocaine on an epidural anesthesia model was carried out on chinchilla rabbits weighing 2.0-2.5 kg. For this, a skin site was projected in the projection of 3-5 lumbar vertebrae and treated with ethanol. Anesthesia of the skin and subcutaneous tissue was performed with 0.25% novocaine solution. In the region of 3-4 lumbar vertebrae, a VUGON TUOHY-PERIDUR needle punctured the epidural space. Compound I and lidocaine in the form of a 2% solution were introduced into the epidural space in the volume calculated by the formula
V =

0.7
The results were taken into account by the absence of deep pain sensitivity and motor activity in the corresponding muscle groups in the area of action of epidural anesthesia (see Table 2).

 Example 6. The antiarrhythmic activity of compound I and lidocaine was studied using a model of calcium chloride arrhythmia in outbred white rats weighing 120-160 g. Animals were anesthetized with sodium ethamine at a dose of 60 mg / kg. The test substances were slowly administered as a single solution intravenously. After 2 minutes, a 10% calcium chloride solution was administered intravenously at a rate of 2 ml / kg. Electrocardiograms were recorded using a KES 01 cardioencephaloscope and a H3021 high-speed recording device. The results of the antiarrhythmic action of compound I and lidocaine were evaluated in percent of surviving animals and in time before ventricular fibrillation occurred (see Table 3).

 PRI me R 7. The study of the local irritating effect of compound I was carried out on rabbits of the chinchilla breed. A 1% solution of compound I was applied once to the conjunctiva of the eye. Observation was carried out for 6 hours every 30 minutes. Conjunctival and scleral vascular injection was not observed, i.e. Compound I has no local irritant effect.

 Example 8. The study of the effect of compound I and lidocaine on the intensity of membrane lipid peroxidation (POL) and mitochondrial energy was performed on female Wistar rats weighing 200-250 g. Mitochondria were isolated simultaneously with microsomes by the method of differentiated centrifugation according to Komat modifications of V.V. Lemeshko.

The study of LPO intensity was carried out in the microsomal fraction of rat liver homogenate. The intensity of the NADP-dependent POL of microsome membranes was determined in a medium of the following composition: 100 mM Tris-HCl buffer (pH 7.4), 1 mM NADP, 4 mM ADP, 12 μM Mohr salt. For ascorbate-dependent lipid peroxidation, the medium contained 100 mM Tris-HCl buffer (pH 7.4), 0.5 mM ascorbate, and 12 μM Mohr salt. In some experiments, an additional 4 mM ADP was added to the incubation medium. The concentration of microcross proteins in the incubation mixture was 0.5-0.6 mg per 1 ml. The incubation mixture was continuously purged with a warmed to 37 ^{° C} air to provide the oxygen saturation of the medium and air mixing. The concentration of compound I and lidocaine was determined by reaction with thiobarbituric acid, after precipitation of the protein with trichloroacetic acid. Protein was determined by Lowry in the modification of Miller.

 The results of a study of the effect of compound I and lidocaine on the intensity of enzymatic and non-enzymatic LPO membranes of rat liver microsomes are presented in Table 4.

 It was noted that the introduction of compound I into the incubation medium leads to a significant inhibition of enzymatic and non-enzymatic LPO. In this case, the effectiveness of compound I with non-enzymatic lipid peroxidation was higher than with enzymatic lipid peroxidation. 50% inhibition (calculated graphically) of non-enzymatic and enzymatic LPO was observed at a concentration of compound I of 0.015 mg / ml (15 mg / L) and 0.195 mg / ml (195 mg / L), respectively.

 Lidocaine with non-enzymatic lipid peroxidation even at a concentration of 2 mg / ml (2 g / l) did not significantly reduce the intensity of free radical lipid oxidation. With enzymatic lipid peroxidation, lidocaine reduced the accumulation of malondialdehyde (MDA) by 50% at a concentration of 1 mg / ml (1 g / l), which is probably due to the inhibitory effect of the NADP-dependent redox chain of microsomes.

 The data obtained indicate that compound I, in contrast to lidocaine, exhibits a pronounced antioxidant effect.

Mitochondrial respiration was measured polarographically using a closed oxygen electrode, recording the curves of oxygen loss on the tape recorder KSP-4. The composition of the reaction medium: 100 mm sucrose, 75 mm KCl, 10 mm KH ₂ PO ₄ , 2 mm MgCl ₂ , 10 mm Tris-HCl buffer (pH 7.4). The oxidation substrate is glutamate + malate (5 ± 5 mM). ADP was added at a concentration of 200 μM, EDTA 0.5 mm.

The respiration rate in metabolic states 2 according to Lardi and Welman (4 according to Chance, v ₂ ), 3 according to Chance (v ₃ ), and in a disconnected state (v _3p ) using 2,4-di-nitrophenol ( DNF) at a concentration of 10 ^-4 M.

 Protein in the samples was determined by Lowry in the modification of Miller.

 The data obtained (see Table 5) indicate that the addition of 0.125-0.25 mg / ml to the cell of compound I leads to a pronounced acceleration of oxygen consumption in state 4 according to Chance. A further increase in the content of this substance in the cell led to a sharp inhibition of mitochondrial respiration.

Noteworthy is also the very significant inhibition of respiration in state 3 (i.e., in the presence of ADP and fn); which, in principle, may indicate a slowdown in the rate of ATP synthesis due to a decrease in the activity of ATP synthetase or an adenosine nucleotide transporter. But since the addition of a disconnector to the cell (state 3 _p ) does not increase the intensity of oxygen consumption by mitochondria, the observed phenomenon is probably not associated with inhibition of mitochondrial ATP synthetase, but is due to inhibition of the respiratory chain. When 0.5 mg / ml of compound I was introduced into the cell, this inhibition was maximum.

 Comparison of the effect on the mitochondrial energy of the cell of compound I and lidocaine indicates that lidocaine in the studied concentrations (up to 2 mg / ml) did not have a significant uncoupling effect, but inhibited the respiratory chain activity, and the maximum effect was observed when 2 mg / ml was introduced into the cell the specified agent. Therefore, the data presented indicate that compound I has a much more pronounced effect on the mitochondrial energy of the cell than lidocaine, which explains its more pronounced antimicrobial and fungicidal effect.

 EXAMPLE 9. The antimicrobial activity of compound I and comparison preparations was studied using the diffusion method in nutrient agar and the method of serial dilutions in a liquid nutrient medium. For bacterial cultures, meat-peptone broth was used; for yeast-like fungus, Saburo agarized and liquid medium was used.

 A suspension of microbial cultures was prepared in physiological saline. The cell concentration in 1 ml was determined using the optical turbidity standard obtained in GNIISiKMEP them. L.A. Tarasevich.

10 ml nutrient agar was poured into sterile Petri dishes. Stainless steel cylinders with a diameter of 8 mm were placed on frozen agar plates. Then, 20 ml of nutrient agar melted and cooled to 40 ^° C was contaminated with a suspension of cells of the test culture under study to a concentration of 10 ⁵ cells / ml. Distributing the culture suspension evenly throughout the entire volume of agar, it was quickly poured onto the frozen first layer. After solidification of the contaminated agar (second layer), cylinders were removed from it. 0.1 ml of the solutions of the test compounds were added to the resulting wells using a concentration of 100 mg / ml. The plates were left for 2 hours at room temperature, then incubated for 48 hours at 37 ^C. The antimicrobial activity was evaluated by the value of growth inhibition zone test culture.

 When using the serial dilution method for each test culture studied, 6 sterile tubes are placed in a tripod. Then prepare a working solution of each compound in meat and peptone broth, a concentration of 1 mg / ml. 4 ml of the working solution are added to the first test tube, and 2 ml of meat and peptone broth are added to the rest. 2 ml of the solution are transferred from the first tube with a sterile pipette into the second and mixed well. Then, by rolling, transfer 2 ml to the next tube and so on, each time using a new pipette. From the fifth tube, 2 ml of the solution was poured. The sixth test tube served as a control.

0.2 ml of cell suspension was added to each tube using a concentration of 10 ⁷ cells / ml. Thus, the concentration of microbial cells in the tested solutions was 10 ⁶ cells / ml. All tubes were incubated in a thermostat at ³⁷ ° C for 48 hours. The antimicrobial activity was evaluated by the presence and intensity rota test culture, comparing with the control tubes.

 The studies performed allowed us to establish a high antimicrobial activity of compound I in relation to Staphylococcus aureus, Escherichia coli, spore-forming saprophytic bacillus, Proteus and yeast-like pathogenic fungus (see Table 6).

 Thus, the compound of formula I in terms of anesthetic, antiarrhythmic, antioxidant, antimicrobial and fungicidal activity significantly surpasses comparison drugs, practically not inferior to them in toxicity, which suggests the possibility of its use in practical medicine, in particular in surgery.

Claims (1)

 DIETHYLAMINOETHYLAMIDE 1-PROPYL-2-OXO-4-HYDROXYCHINOLIN-3-CARBONIC ACID HYDROCHLORIDE, MANIFESTING ANESTHESIS, ANTIVARISTINUMINIDINITINUYTINUINTINUINIDINUINIDINUTINU 1-propyl-2-oxo-4-hydroxyquinoline-3-carboxylic acid diethylaminoethylamide hydrochloride of the formula

showing anesthetic, antiarrhythmic, antioxidant, antimicrobial and fungicidal activity.

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